ABSTRACT Studies of the microbiome of older individuals has shown greatly reduced numbers of known beneficial bacteria coinciding with a greater presence of potentially detrimental bacterial strains. Not surprisingly, the decrease in beneficial microbes coincides with a loss of anti-inflammatory activity and genes associated with short-chain fatty acid production. This age-related decline in the gut microbiome is also an important determinant of age-associated pathological states including neurodegeneration, chronic inflammation, frailty, cognitive decline, type 1 and type 2 diabetes, musculoskeletal degeneration, as well as nonalcoholic fatty liver disease, cardiovascular disease, and development of certain types of cancer. The need to increase overall consumption of resistant starch (RS) prebiotics in the elderly population is clear. The difficulty arises in providing RS in sufficient quantities (20?30g/daily) to these individuals in a form that is easily stored and readily consumed. In this project, Design-Zyme LLC will 1) Develop a rapid assay for evaluation of prebiotic nutrient component efficacy on known human gut microbes, and 2) Optimize a safe, digestion-resistant molecule specifically targeting beneficial human gut microbiota deficiencies in the elderly. To accomplish this, we will design an assay to quickly and easily evaluate the growth of common strains of beneficial gut bacteria and then use this assay to screen individual molecular components of a known resistant starch prebiotic compound. At the end of Phase I, we will have developed an easy-to-use assay for determining effectiveness of individual prebiotics. Furthermore, using this assay, we will have identified specific molecules that promote the proliferation of beneficial gut bacteria, especially those in the aging and elderly populations. During Phase II of this proposal, we will establish how to best modify and formulate the molecules from Phase I to resist breakdown in solution while concomitantly allowing for more concentrated solutions to be manufactured. At the end of Phase II, we will have established how to protect these molecules from breakdown in a water solution, allowing for the creation of pre-mixed aqueous beverages capable of retaining their prebiotic functionality with a minimum shelf-life of six months. Research into prebiotic nutrient compounds remains an important, emerging field. The techniques and assays developed in this grant proposal will be widely applicable to a range of prebiotic compounds and will advance the development of effective, concentrated prebiotics capable of directly modulating the human gut microbiome and improving overall health in the elderly population.